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S. C. Santos, C. A. Custódio, J. F. Mano, Human Protein‐Based Porous Scaffolds as Platforms for Xeno‐Free 3D Cell Culture, Advanced Healthcare Materials, 2022.


Abstract:

Extracellular matrix and protein-based biomaterials emerge as attractive sources to produce scaffolds due to their great properties regarding biocompatibility and bioactivity. In addition, there are concerns regarding the use of animal-derived supplements in cell culture not only due to the risk of transmission of xenogeneic contaminants and antigens but also due to ethical issues associated with collection methods. Herein, a novel human protein-derived porous scaffold produced from platelet lysates (PL) as platform for xeno-free 3D cell culture has been proposed. Human PL are chemically modified with methacryloyl groups (PLMA) to make them photocrosslinkable and used as precursor material to produce PLMA-based sponges. The herein reported human-based sponges have highly tunable morphology and mechanical properties, with an internal porous structure and Young’s modulus dependent on the concentration of the polymer. Human adipose-derived stem cells (hASCs) are cultured on top of PLMA sponges to validate their use for 3D cell culture in xeno-free conditions. After 14 days hASCs remained viable, and results show that cells are able to proliferate during time even in the absence of animal-derived supplementation. This study reveals for the first time that such scaffolds can be promising platforms for culture of human cells avoiding the use of any animal-derived supplement.


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C. F. Monteiro, C. A. Custódio, J. F. Mano, Bioengineering a humanized 3D tri-culture osteosarcoma model to assess tumor invasiveness and therapy response, Acta Biomaterialia, 2021.


Abstract:

To date, anticancer therapies with evidenced efficacy in preclinical models fail during clinical trials. The shortage of robust drug screening platforms that accurately predict patient’s response underlie these misleading results. To provide a reliable platform for tumor drug discovery, we herein propose a relevant humanized 3D osteosarcoma (OS) model exploring the potential of methacryloyl platelet lysates (PLMA)-based hydrogels to sustain spheroid growth and invasion. The architecture and synergistic cell-microenvironment interaction of an invading tumor was recapitulated encapsulating spheroids in PLMA hydrogels, alone or co-cultured with osteoblasts and mesenchymal stem cells. The stem cells alignment toward OS spheroid suggested that tumor cells chemotactically attracted the surrounding stromal cells, which supported tumor growth and invasion into the hydrogels. The exposure of established models to doxorubicin revealed an improved drug resistance of PLMA-based models, comparing with scaffold-free spheroids. The proposed OS models highlighted the feasibility of PLMA hydrogels to support tumor invasion and recapitulate tumor-stromal cell crosstalk, demonstrating the potential of this 3D platform for complex tumor modelling. STATEMENT OF SIGNIFICANCE: Cell invasion mechanisms involved in tumor progression have been recapitulated in the field of 3D in vitro modeling, leveraging the great advance in biomimetic materials. In line with the growing interest in human-derived biomaterials, the aim of this study is to explore for the first time the potential of methacryloyl platelet lysates (PLMA)-based hydrogels to develop a humanized 3D osteosarcoma model to assess tumor invasiveness and drug sensitivity. By co-culturing tumor spheroids with human osteoblasts and human mesenchymal stem cells, this study demonstrated the importance of the synergistic tumor cell-microenvironment interaction in tumor growth, invasion and drug resistance. The established 3D osteosarcoma model highlighted the feasibility of PLMA hydrogels as a relevant 3D platform for complex tumor modelling


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C. F. Monteiro, S. C. Santos, C. A. Custódio, J. F. Mano, Human Platelet Lysates-Based Hydrogels: A Novel Personalized 3D Platform for Spheroid Invasion Assessment, Advanced Science, 2020.


Abstract:

Fundamental physiologic and pathologic phenomena such as wound healing and cancer metastasis are typically associated with the migration of cells through adjacent extracellular matrix. In recent years, advances in biomimetic materials have supported the progress in 3D cell culture and provided biomedical tools for the development of models to study spheroid invasiveness. Despite this, the exceptional biochemical and biomechanical properties of human-derived materials are poorly explored. Human methacryloyl platelet lysates (PLMA)-based hydrogels are herein proposed as reliable 3D platforms to sustain in vivo-like cell invasion mechanisms. A systematic analysis of spheroid viability, size, and invasiveness is performed in three biomimetic materials: PLMA hydrogels at three different concentrations, poly(ethylene glycol) diacrylate, and Matrigel. Results demonstrate that PLMA hydrogels perfectly support the recapitulation of the tumor invasion behavior of cancer cell lines (MG-63, SaOS-2, and A549) and human bone-marrow mesenchymal stem cell spheroids. The distinct invasiveness ability of each cell type is reflected in the PLMA hydrogels and, furthermore, different mechanical properties produce an altered invasive behavior. The herein presented human PLMA-based hydrogels could represent an opportunity to develop accurate cell invasiveness models and open up new possibilities for humanized and personalized high-throughput screening and validation of anticancer drugs.


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S. C. Santos, C. A. Custódio, J. F. Mano, Photopolymerizable Platelet Lysate Hydrogels for Customizable 3D Cell Culture Platforms, Advanced Healthcare Materials, 2018.


Abstract:

3D cell culture platforms have emerged as a setting that resembles in vivo environments replacing the traditional 2D platforms. Over the recent years, an extensive effort has been made on the development of more physiologically relevant 3D cell culture platforms. Extracellular matrix–based materials have been reported as a bioactive and biocompatible support for cell culture. For example, human plasma derivatives have been extensively used in cell culture. Despite all the promising results, in most cases these types of materials have poor mechanical properties and poor stability in vitro. Here plasma-based hydrogels with increased stability are proposed. Platelet lysates are modified by addition of methacryloyl groups (PLMA) that polymerize in controlled geometries upon UV light exposure. The hydrogels could also generate porous scaffolds after lyophilization. The results show that PLMA materials have increased mechanical properties that can be easily adjusted by changing PLMA concentration or modification degree. Cells readily adhere, proliferate, and migrate, exhibiting high viability when encapsulated in PLMA hydrogels. The innovation potential of PLMA materials is based on the fact that it is a complete xeno-free solution for human cell culture, thus an effective alternative to the current gold standards for 3D cell culture based on animal products.


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